Wavelength scanning apparatus and method of use thereof

ABSTRACT

A wavelength scanning apparatus that detects at least four different fluorescent emission wavelengths simultaneously or nearly simultaneously is described. The wavelength scanning apparatus includes a heating block having at least four sample wells, each sample well configured for receiving a sample, at least four excitation activation apertures, and at least four fluorescence emission discharge apertures. The wavelength scanning apparatus also includes an analysis scanner having at least four light sources, where the at least four light sources excite at least four fluorophores, at least four excitation light filters that filter out light except that of the desired excitation wavelength/s, at least four fluorescence emission light filters that filter out light except that of the desired fluorescent emission wavelengths, and at least four photodetectors to detect light of the desired fluorescent emission wavelengths.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims priority to and is a Continuation-in-Part ofU.S. patent application Ser. No. 14/809,175, filed on Jul. 25, 2015,which is expressly incorporated herein by reference in its entirety.

BACKGROUND

Fluorescence detection of nucleic acids and proteins is carried out by avariety of apparatuses and methods, including capillary electrophoresis,deoxyribonucleic acid (DNA) sequencing with fluorescent dyes, andmicrofluidic fluorescence detection. Methods and apparatus forfluorescence detection of nucleic acids and proteins generally includefour common elements: a light source for excitation of fluorophores, afluorophore capable of excitation and emission, filters to isolate awavelength emitted from an excited fluorophore, and a detector thatdetects the emitted wavelength from the excited fluorophore and producesan electrically recordable output.

When methods and apparatus of fluorescence detection are used fornucleic acid detection, such methods may require polymerase chainreaction (PCR) or isothermal amplification to obtain the desired outputsignal. The fluorescence detection apparatus generally includes aheating block having one or more sample wells configured for receivingvessels where PCR or isothermal amplification may take place. Ininstances where the heating block has at least two wells, a movablescanning component may be necessary where either the heating block orthe detector is moved in order to measure the fluorescence of a samplein each of the different sample wells. Typically, the movable scanningcomponent contains dichroic mirrors, filter wheels, and photomultipliertubes to direct, isolate, and convert the fluorescence emissions fromthe samples to an electric output. These components are costly and limitthe simultaneous detection of multiple wavelengths. Detection of asingle fluorescent emission wavelength increases the time required formeasuring fluorescent emission wavelengths from multiple sample wells,thereby decreasing efficiency and increasing the time required tocomplete the analysis of multiple sample wells.

It is desirable to eliminate expensive parts from the movable scanningcomponent used in fluorescence detection. It is also desirable toprovide a fluorescence detection system capable of detecting at leastfour fluorescent emission wavelength emissions simultaneously or nearlysimultaneously.

SUMMARY

An apparatus for fluorescence detection through a wavelength scanningapparatus is described. A wavelength scanning apparatus usingfluorescence emissions to test for the presence of at least four nucleicacid sequences or proteins simultaneously or nearly simultaneously alsois described.

A nucleic acid analysis method for performing fluorescence detection ofmultiple fluorescence emission wavelengths simultaneously or nearlysimultaneously is also described.

A DNA analysis method for performing fluorescence detection of multiplefluorescence emission wavelengths simultaneously or nearlysimultaneously is also described.

The following detailed description is exemplary and explanatory only andis not necessarily restrictive of the invention as claimed. Theaccompanying drawings, which are incorporated in and constitute a partof the specification, illustrate embodiments of the invention andtogether with the detailed description, serve to explain the principlesof the invention.

BRIEF DESCRIPTION OF THE DRAWINGS

The numerous advantages of the present invention may be betterunderstood by those skilled in the art by reference to the accompanyingfigures.

FIG. 1 represents a wavelength scanning apparatus.

FIG. 2A is a vertical cross sectional view of a heating block of thewavelength scanning apparatus.

FIG. 2B is a vertical cross section view of a well of the wavelengthscanning apparatus.

FIG. 3A is a front angled view of a analysis scanner of the wavelengthscanning apparatus.

FIG. 3B is a back angled view of a analysis scanner of the wavelengthscanning.

FIG. 4 depicts a heating block of a wavelength scanning apparatus.

FIG. 5 illustrates a method for testing for the presence of nucleic acidsamples using the wavelength scanning apparatus.

FIG. 6 illustrates alignment of an analysis scanner with a plurality ofsample wells.

FIG. 7 represents another wavelength scanning apparatus.

FIG. 8A is a vertical cross sectional view of a heating block of thewavelength scanning apparatus.

FIG. 8B is a vertical cross section view of a well of the wavelengthscanning apparatus.

FIG. 9 is a heating block of a wavelength scanning apparatus.

FIG. 10 illustrates a method for testing for the presence of nucleicacid samples using the wavelength scanning apparatus with a resetbutton.

DETAILED DESCRIPTION

A wavelength scanning apparatus that detects at least four fluorescentemission wavelengths simultaneously or nearly simultaneously isdescribed. The wavelength scanning apparatus includes a heating blockhaving at least four sample wells, where each sample well is configuredfor receiving a sample, at least four excitation activation apertures,and at least four fluorescence emission discharge apertures. Theexcitation activation apertures and fluorescence emission dischargeapertures are arranged in a right angle or nearly a right angle to eachother for fluorescence detection. The wavelength scanning apparatus alsoincludes an analysis scanner having at least four light sources, whereeach of the at least four light sources excites a different fluorophore,at least four excitation light filters that filter out light except thatof the desired excitation wavelength/s, at least four fluorescenceemission light filters that filter out light except that of the desiredfluorescent emission wavelength/s, and at least four photodetectors todetect light of the desired fluorescent emission wavelengths. Each ofthe at least four light sources may be paired with a differentexcitation light filter and a different fluorescent emission lightfilter. The scanner may remain stationary along one face of the heatingblock for detection of at least four fluorescent emission wavelengthssimultaneously or nearly simultaneously. The analysis scanner may movelaterally along one face of the heating block to a plurality ofpredetermined locations for detection of at least four differentfluorescent emission wavelengths from each sample. Thus, while theanalysis scanner may simultaneously or nearly simultaneously analyze atleast four different fluorescent emission wavelengths, the analysisscanner is analyzing one fluorescent emission wavelength from a singlesample well at a time. The movement of the analysis scanner laterallyalong one face of the heating block may be constrained by threadedmovement.

FIG. 1 represents a wavelength scanning apparatus 400. The wavelengthscanning apparatus 400 comprises a heating block 100, an analysisscanner 300, a stepper motor 401, a stepper screw 402, a stepper screwhole 111, and a structural support 403 having a guide rail 405. Theheating block 100 is attached to the structural support 403 via theheating block screw holes 202 such that the heating block 100 issuspended horizontally by the structural support 403. The analysisscanner 300 may be attached to the structural support 403 by insertionof the guide rail 405 through the guide rail hole 112 so that theanalysis scanner is suspended horizontally such that the flat surface104 of the analysis scanner 300 aligns with the bottom of the heatingblock 100, and the front wall 107 of the analysis scanner 300 alignswith the front of the heating block 100. The bottom of the heating block100 may nearly contact the flat surface 104 of the analysis scanner 300,and the front of the heating block 100 may nearly contact the front wall107 of the analysis scanner 300. The bottom of the heating block 100 maycontact the flat surface 104 of the analysis scanner 300, and the frontof the heating block 100 may contact the front wall 107 of the analysisscanner 300, where such contact by the flat surface 104 and the frontwall 107 allows for lateral movement of the analysis scanner 300. Thestepper motor 401 may be attached to the structural support 403 suchthat it is suspended horizontally below the heating block 100. Thestepper screw 402 is connected to the stepper motor 401. The structuralsupport 403 may include a rectangular piece of material (e.g. plastic ormetal) (not shown) that serves as a base platform and has one or morevertical supports configured for attachment to the heating block 100 viathe heating block screw holes, the stepper motor 401, and the guide rail405. The structural support may include a casing (not shown) configuredto enclose the wavelength scanning apparatus 400. In embodiments thecasing has a lid or a flap that opens above the heating block 100configured for introduction of DNA samples into the heating block 100.The casing may be made of any material consistent with the desiredoperation of the device, such as plastic or aluminum.

The stepper motor 401 is configured for turning the stepper screw 402 apredetermined distance wherein a plurality of excitation apertures 105of the analysis scanner 300 align with a plurality of excitationactivation apertures 102 of the heating block 100, and a plurality offluorescence emission discharge apertures 103 of the heating block 100align with a plurality of emission apertures 108 of the analysis scanner300 (e.g. the geometric centers of these corresponding aperturessubstantially align, thus sufficiently align for transmission of thedesired excitation and emission wavelengths). For example, theexcitation aperture 105 substantially aligns with the excitationactivation aperture 102 and the emission aperture 108 substantiallyaligns with the fluorescence emission discharge aperture 103. Theanalysis scanner 300 may move laterally in either direction horizontallyalong the stepper screw 402 via the stepper motor 401 rotating thestepper screw 402 clockwise or counter-clockwise. The stepper screw 402and the stepper screw hole 111 may be threaded where the movement of theanalysis scanner 300 by the stepper motor 401 is constrained by athreaded movement.

The stepper motor 401, the light sources, and the photodetectors of theanalysis scanner 300 may be regulated (e.g. turned on and turned off) bya controller. The controller may be configured to move the analysisscanner 300 backward and forward along the heating block 100 in apredetermined amount via the stepper screw 402, to turn the lightsources on and off, to turn the photodetectors on and off, to turn theheating element 404 of the heating block 100 on and off, and to resetthe analysis scanner. For example, the controller may regulate thestepper motor, light sources, and photodetectors in a sequence orcombination. A computer program may be used to configure the controller.

FIG. 7 represents a wavelength scanning apparatus. The wavelengthscanning apparatus 400 includes a heating block 100, an analysis scanner300, a stepper motor 401, a stepper screw 402, a stepper screw hole 111,and a structural support 403 having a guide rail 405 as described inFIG. 1. The wavelength scanning apparatus 400 also includes a resetbutton 410, a reset trigger 411, at least one stepper nut screw 412, anda stepper nut 413. The reset button 410 is in mechanical communicationwith the structural support 403. The reset trigger 411 is in mechanicalcommunication with the reset button 410 and the structural support 403.The reset button 410 and the reset trigger 411 are configured to alignthe analysis scanner 300 with the florescence emission dischargeaperture 103 prior to and/or after fluorescence detection analysis. Thereset button 410 may be in electronic communication with a controller,such that the controller may initiate a reset of the analysis scannerafter each fluorescence detection analysis. The reset of the analysisscanner 300 provides that the analysis scanner 300 is in the samealignment with the florescence emission discharge aperture 103 for eachfluorescence detection analysis.

The stepper nut screw 412 is in mechanical communication with theanalysis scanner 300 and the stepper screw 402. The stepper nut 413 isin mechanical communication with the stepper nut screw 412 and thestepper screw 402. The stepper nut screw 412 and the stepper nut 413 areconfigured for securing the analysis scanner to the stepper screw 402.

FIG. 2A is a vertical cross sectional view of the heating block 100.Sample wells 101 have a shape configured to hold a sample, for example,a 0.2 ml PCR tube. The excitation activation aperture 102 may be, forexample, 2 mm in diameter and cylindrical in shape. The fluorescenceemission discharge aperture 103 may be, for example, cone shaped with adiameter of 2 mm at the back and 1 mm at the front.

FIG. 2B shows a vertical cross sectional view of the sample well 101.The sample well 101 has a shape configured to hold a sample, forexample, a 0.2 ml PCR tube. The excitation activation aperture 102 maybe, for example, 2 mm in diameter and cylindrical in shape. Thefluorescence emission discharge aperture 103 may be, for example, coneshaped with a diameter of 2 mm at the back and 1 mm at the front.

FIG. 8A is a vertical cross sectional view of the heating block 100. Thefluorescence emission discharge aperture 103 may be cylindrical in shapewith a diameter of 2 mm. The fluorescence emission discharge aperturemay be from 0.5 mm to 3.0 mm from the top of the excitation activationaperture 103. The front of the heating block 100 may be lower than theback of the heating block 100, such that there is angled downward slopefrom the back to the front of the heating block 100. The front of theheating block 100 may be 4 mm higher than the back of the heating block100.

FIG. 8B shows a vertical cross sectional view of the sample well 101.The fluorescence emission discharge aperture 103 may be cylindrical inshape with a diameter of 2 mm. The fluorescence emission dischargeaperture may be from 0.5 mm to 3.0 mm from the top of the excitationactivation aperture 103.

FIGS. 3A and 3B represent a front side angled view and a back sideangled view, respectively, of the analysis scanner 300 of the wavelengthscanning apparatus. The analysis scanner 300 includes a flat surface104, a front wall 107, a side surface 110, at least four excitationapertures 105, at least four emission apertures 108, at least fourexcitation light filters 130, at least four fluorescence emission lightfilters 120, an excitation recessed area 113, an emission recessed area114, at least four light sources 150, and at least four photo detectors140. The at least four light sources may originate from a single lightsource that is split by way of light channels, light pipes, and the liketo provide the at least four light sources, or from multiple lightsources.

The analysis scanner 300 has dimensions that correspond with dimensionsof and substantial alignment of apertures with the heating block 100.Such dimension correspondence is explained further in paragraphs thatfollow in relation to the operation of the wavelength scanning apparatus400.

The analysis scanner 300 has a stepping block shape, where a front ofthe analysis scanner 300 has a step indentation formed by the flatsurface 104 being substantially perpendicular with the front wall 107along a top side of the analysis scanner 300. For example, the flatsurface 104 extends toward the back of the analysis scanner 300 untilthe flat surface 104 meets the front wall 107 in a perpendicular manner,such that the step indentation forms an approximate right angle. Theflat surface 104 may have a depth of 10.5 mm, and the front wall 107 mayhave a height of 9.5 mm.

The analysis scanner 300 may have an excitation aperture 105 having atop and a bottom. The excitation aperture 105 extends from the flatsurface 104 downward to the excitation recessed area 113 at the bottomof the analysis scanner 300. The excitation aperture 105 may becylindrical with a diameter of 6.45 mm at the bottom and 4 mm at thetop. The analysis scanner 300 may have an emission aperture 108 having afront and a back. The emission aperture 108 extends from the front wall107 backward to the emission recessed area 114 at the back of theanalysis scanner 300. The emission aperture 108 may be cone shapedhaving a diameter of 4 mm at the front and 6.45 mm at the back. Theemission aperture 108 may be cylindrical having a diameter of 2.5 mm.The analysis scanner 300 may have an excitation light filter 130. Theexcitation light filter 130 may be set inside the excitation aperture105 toward the flat surface 104 and configured for filtering outwavelengths of light except that of an excitation wavelength. Forexample, a first excitation light filter is configured to filter lightexcept light of the absorption wavelength of a first fluorophore (e.g.494 nm, 500 nm), a second excitation light filter is configured tofilter light except light of the absorption wavelength of a secondflurophore (e.g. 515 nm, 564 nm), a third excitation light filter isconfigured to filter light except light of the absorption wavelength ofa third fluorophore (e.g. 559 nm, 645 nm), and a fourth excitation lightfilter is configured to filter light expect light of the absorptionwavelength of a fourth fluorophore (e.g. 647 nm, 747 nm).

The analysis scanner 300 may have a fluorescence emission light filter120. The fluorescence emission light filter 120 may be set inside theemission aperture 108 toward the front wall 107 and configured forfiltering out wavelengths of light except that of a fluorescent emissionwavelength. For example, a first fluorescence light filter is configuredto filter light except light of the fluorescent emission wavelength ofthe first fluorophore (e.g. 521 nm, 520 nm), a second fluorescence lightfilter is configured to filter light except light of the fluorescentemission wavelength of the second fluorophore (e.g. 650 nm, 590 nm), athird fluorescence light filter is configured to filter light exceptlight of the fluorescent emission wavelength of the third fluorophore(e.g. 578 nm, 671 nm), and a fourth fluorescence light filter isconfigured to filter light except light of the fluorescent emissionwavelength of the fourth fluorophore (e.g. 670 nm, 776 nm).

The scanner 300 has a plurality of light sources 150 wherein a firstlight source is configured to emit light of a first wavelength to excitethe first fluorophore corresponding to a first nucleic acid primer (e.g.DNA primer, RNA primer), a second light source is configured to emitlight of a second wavelength to excite the second fluorophorecorresponding to a second nucleic acid primer, a third light source isconfigured to emit light of a third wavelength to excite the thirdfluorophore corresponding to a third nucleic acid primer, and a fourthlight source is configured to emit light of a fourth wavelength toexcite the fourth fluorophore corresponding to a fourth nucleic acidprimer. The scanner 300 contains a plurality of photodetectors 140configured to detect fluorescent emission wavelengths from the pluralityof fluorophores. For example, a first photodetector is configured todetect the fluorescent emission wavelength from the first fluorophore, asecond photodetector is configured to detect the fluorescent emissionwavelength from the second fluorophore, a third photodetector isconfigured to detect the fluorescent emission wavelength from the thirdfluorophore, and a fourth photodetector is configured to detect thefluorescent emission wavelength from the fourth fluorophore. Theplurality of photodetectors 140 may be photodiodes that detect thephotons of the fluorescent emission wavelength.

Referring to FIG. 3B the excitation recessed area 113 is a recessed areaon the bottom of the analysis scanner 300. The excitation recessed area113 may be rectangular in shape and have dimensions that correspond withthe dimensions of the analysis scanner 300. For example, the excitationrecessed area 113 may be 45.5 mm length, 21.14 mm height, and 6 mmdepth. The excitation recessed area 113 may be sized for receiving anexcitation printed circuit board. The excitation printed circuit boardincludes a plurality of light sources. The light sources preferably arelight emitting diodes (LEDs) capable of emitting light at a plurality ofdifferent excitation wavelengths. Each light source is of slightlysmaller diameter than the diameter of the excitation aperture 105 suchthat when the circuit board is received by the excitation recessed area113, each light source fits within the corresponding excitation aperture105. The excitation printed circuit board may be secured to theexcitation recessed area 113 by screwing the circuit board into theanalysis scanner 300 at a plurality of excitation circuit holes 302.

Referring to FIG. 3B the emission recessed area 114 is a recessed areaon the back of the analysis scanner 300. The emission recessed area 114may be rectangular in shape and has dimensions that correspond with thedimensions of the analysis scanner 300. For example, the emissionrecessed area 114 may be a height of 18 mm, a length of 45.5 mm, and adepth of 6 mm. The emission recessed area 114 may be configured forreceiving an emission printed circuit board. The emission printedcircuit board includes at least four photodetectors. Each photodetectorpreferably is a photodiode configured for detecting at least onefluorescent emission wavelength. The photodetector is of slightlysmaller diameter than the diameter of the emission aperture 108 suchthat when the circuit board is received by the emission recessed area114, each photodetector fits within the corresponding emission aperture108. The circuit board may be secured to the emission recessed area 114by screwing the circuit board into the analysis scanner 300 at aplurality of emission circuit holes 303.

Referring to FIG. 3A and FIG. 3B the analysis scanner 300 may have astepper screw hole 111 and a guide rail hole 112. The stepper screw hole111 may be an aperture extending horizontally through the analysisscanner 300 and is configured for receiving a stepper screw 402. Thestepper screw hole 111 may be a variety of lengths corresponding to thelength of the analysis scanner 300; for example, the screw hole 111 maybe 46.5 mm in length. The stepper screw hole 111 may be threaded wherethe movement of the analysis scanner 300 along the stepper screw 402 isconstrained by the threads of the stepper screw hole 111 and the stepperscrew 402. The stepper screw hole 111 may be unthreaded when themovement of the analysis scanner 300 along the stepper screw hole 402 isconstrained by at least one stepper nut screw 412 and a stepper nut 413.The guide rail hole 112 is an aperture extending horizontally throughthe analysis scanner 300 and is configured to receive a guide rail. Theguide rail hole 112 may be a variety of lengths corresponding to thelength of the analysis scanner 300; for example, the guide rail hole maybe 46.5 mm in length. In embodiments, the guide rail may be connected tothe structural support 403 and may be configured to hold the analysisscanner 300 in place within the wavelength scanning apparatus 400.

FIG. 4 depicts a heating block 100 of the wavelength scanning apparatus400. The heating block 100 includes a top side 210, a bottom side 240, afront side 220, and a back side 230. The heating block 100 includes atleast four sample wells 101, at least four excitation activationapertures 102, and at least four fluorescence emission dischargeapertures 103. The heating block 100 may be of any metal, ceramic, orother heat resistant material, such as aluminum. The heating block 100may be of any dimension, such as a t-block shape having a first lengthof 166 millimeters (mm) in length, a second length of 100 mm, a width of12 mm, a first height of 20 mm, and a second height of 8.41 to 9.63 mm.The front 220 of the heating block 100 may be lower than the back 230 ofthe heating block 100, such that there is angled downward slope from theback to the front of the heating block 100. The front of the heatingblock 100 may be 4 mm higher than the back of the heating block 100. Thesample well 101 may be formed as part of the heating block 100, wherethe sample well 101 is a hollowed out portion of the heating block 100,having a bottom and a top. The sample well 101 may extend from the topside 210 of the heating block 100 toward the bottom side 240 of theheating block 100 where the sample well 101 terminates in the excitationactivation aperture 102. The sample well 101 may be of any shape anddiameter compatible with sample retention, entry of the excitationwavelength, and exit of the fluorescent emission wavelength (e.g.cylindrical, rectangular prism, and the like). The sample well 101 maybe of tapered cylindrical shape configured for fitting a 0.2 milliliter(ml) PCR tube. The excitation activation aperture 102 may extend fromthe bottom side of the heating block 100 toward the top side of theheating block 100 where the excitation activation aperture 102transitions into the bottom of the sample well 101, thus forming anoutlet the diameter of the excitation activation aperture 102. Theexcitation activation aperture 102 may substantially align with thebottom of the sample well 101 such that the approximate geometric centerof the excitation activation aperture 102 substantially aligns with thegeometric center of the sample well 101. The excitation activationaperture 102 may have a cylindrical shape. The excitation activationaperture 102 may have a 2 millimeter (mm) diameter.

A fluorescence emission discharge aperture 103, having a front and aback, may be formed on the front side 220 of the heating block 100 andextend toward the back side 230 of the heating block 100 where thefluorescence emission discharge aperture 103 transitions into a side ofthe sample well 101, forming an outlet having a diameter substantiallyequal to the back of the fluorescence emission discharge aperture 103,such that the excitation activation aperture 102 and the fluorescenceemission discharge aperture 103 are in light communication via thesample well 101. The fluorescence emission discharge aperture 103 may beperpendicular to the sample well 101, such that an approximate rightangle is formed between the excitation activation aperture 102 and thefluorescence emission discharge aperture 103. The fluorescence emissiondischarge aperture 103 may have a cone shape where the diameter of thefront of the fluorescence emission discharge aperture 103 is smallerthan the diameter of the back of the fluorescence emission dischargeaperture 103. The front of the fluorescence emission discharge aperture103 may be 1 mm in diameter and the back of the fluorescence emissiondischarge aperture 103 may be 2 mm in diameter. The plurality of samplewells, excitation activation apertures and discharge apertures may besubstantially the same as the sample well 101, the excitation activationaperture 102, and the fluorescence emission discharge aperture 103.Other sample well designs for the heating block 100 that are compatiblewith the operating principles of the wavelength scanning apparatus 400may be used.

Referring to FIG. 4, the heating block 100 may have one or more heatsink screw holes 201 and one or more heating block screw holes 202. Theheat sink screw hole 201 may be configured for mounting a heatingelement 404, having a front side and a back side, and a heat sink,having a front side and a back side, to the heating block 100 where thefront side of the heating element 404 contacts the back of the heatingblock 100, and the front side of the heat sink contacts the back side ofthe heating element 404. The heating element 404 may be a Peltierheater, and the heat sink may be a pinned heat sink. The heating blockscrew holes 202 may be configured for attaching the heating block 100 tothe structural support 403. The structural support 403 may be configuredfor supporting the heating block 100 such that the wavelength scanningapparatus 400 may have its top side facing upright (or upward).

FIG. 9 illustrates a heating block 100 of the wavelength scanningapparatus. The heating block 100 includes a top side 210, a bottom side240, a front side 220, a back side 230, at least four sample wells 101,and at least four excitation activation apertures 102 as described inFIG. 4. The heating block 100 may include a fluorescence emissiondischarge aperture 103 that may be cylindrical in shape with a diameterof 2.5 mm. The fluorescence emission discharge aperture may be from 0.5mm to 3.0 mm from the top of the florescence emission excitationactivation aperture 103.

Unlike in FIG. 1, FIG. 2A, and FIG. 4, where the wavelength scanningapparatus 400 is represented with eight of the sample wells 101 in theheating block 100, when the heating block 100 includes four of thesample wells 101 and the analysis scanner 300 includes four sets ofapertures, the analysis scanner 300 may move to a series of positions asrepresented in FIG. 6. Referring to FIG. 6, in a first position 610 afirst excitation activation aperture is substantially aligned with afourth excitation aperture and a first fluorescence emission dischargeaperture is substantially aligned with a fourth emission aperture. Themovement of the analysis scanner 300 occurs via the stepper motor 401wherein the stepper motor 401 is configured to turn the stepper screw402 a predetermined distance to achieve substantial alignment of thefirst position.

The analysis scanner 300 then may move to a second position 620 wherethe first excitation activation aperture is substantially aligned with athird excitation aperture, the first fluorescence emission dischargeaperture is substantially aligned with a third emission aperture, asecond excitation activation aperture is substantially aligned with thefourth excitation aperture, and a second fluorescence emission dischargeaperture is substantially aligned with the fourth emission aperture. Themovement of the analysis scanner 300 occurs via the stepper motor 401where the stepper motor 401 is configured to turn the stepper screw 402a predetermined distance to achieve substantial alignment of the secondposition.

The analysis scanner 300 then may move to a third position 630 where thefirst excitation activation aperture is substantially aligned with asecond excitation aperture, the first fluorescence emission dischargeaperture is substantially aligned with a second emission aperture, thesecond excitation activation aperture is substantially aligned with thethird excitation aperture, the second fluorescence emission dischargeaperture is substantially aligned with the third emission aperture, athird excitation activation aperture is substantially aligned with thefourth excitation aperture, and a third fluorescence emission dischargeaperture is substantially aligned with the fourth emission aperture. Themovement of the analysis scanner 300 occurs via the stepper motor 401where the stepper motor 401 is configured to turn the stepper screw 402a predetermined distance to achieve substantial alignment of the thirdposition.

The analysis scanner 300 then may move to a fourth position 640 wherethe first excitation activation aperture is substantially aligned with afirst excitation aperture, the first fluorescence emission dischargeaperture is substantially aligned with a first emission aperture, thesecond excitation activation aperture is substantially aligned with thesecond excitation aperture, the second fluorescence emission dischargeaperture is substantially aligned with the second emission aperture, thethird excitation activation aperture is substantially aligned with thethird excitation aperture, the third fluorescence emission dischargeaperture is substantially aligned with the third emission aperture, afourth excitation activation aperture is substantially aligned with thefourth excitation aperture, and a fourth fluorescence emission dischargeaperture is substantially aligned with the fourth emission aperture. Themovement of the analysis scanner 300 occurs via the stepper motor 401where the stepper motor 401 is configured to turn the stepper screw 402a predetermined distance to achieve substantial alignment of the fourthposition. In the fourth position, the analysis scanner 300 detects fourfluorescent emission wavelengths simultaneously or nearlysimultaneously.

The analysis scanner 300 then may move to a fifth position 650 where thesecond excitation activation aperture is substantially aligned with thefirst excitation aperture, the second fluorescence emission dischargeaperture is substantially aligned with the first emission aperture, thethird excitation activation aperture is substantially aligned with thesecond excitation aperture, the third fluorescence emission dischargeaperture is substantially aligned with the second emission aperture, thefourth excitation activation aperture is substantially aligned with thethird excitation aperture, and the fourth fluorescence emissiondischarge aperture is substantially aligned with the third emissionaperture. The movement of the analysis scanner 300 occurs via thestepper motor 401 where the stepper motor 401 is configured to turn thestepper screw 402 a predetermined distance to achieve substantialalignment of the fifth position.

The analysis scanner 300 then may be moved to a sixth position 660 wherethe third excitation activation aperture is substantially aligned withthe first excitation aperture, the third fluorescence emission dischargeaperture is substantially aligned with the first emission aperture, thefourth excitation activation aperture is substantially aligned with thesecond excitation aperture, and the second fluorescence emissiondischarge aperture is substantially aligned with the fourth emissionaperture. The movement of the analysis scanner 300 occurs via thestepper motor 401 where the stepper motor 401 is configured to turn thestepper screw 402 a predetermined distance to achieve substantialalignment of the sixth position.

The analysis scanner 300 then may be moved to a seventh position 670where the fourth excitation activation aperture is substantially alignedwith the first excitation apertures and the fourth fluorescence emissiondischarge aperture is substantially aligned with the first emissionaperture. The movement of the analysis scanner 300 occurs via thestepper motor 401 where the stepper motor 401 is configured to turn thestepper screw 402 a predetermined distance to achieve substantialalignment of the seventh position. Upon completion of detection inpositions one through seven, detection of four different fluorescentemission wavelengths has occurred from each of the four different samplewells 101 with different fluorescent emission wavelengths.

A method 500 is used to analyze at least four deoxyribonucleic acid(DNA) samples for at least four different DNA sequences (e.g.nucleotides or oligonucleotides). A DNA sample may contain all, some, ornone of the DNA sequences. Each DNA sample includes a plurality of DNAprimers to detect the DNA sequences that are present in each sample.Each DNA primer could be labeled with a fluorophore, each fluorophorehaving unique absorption and emission properties. Detection of the DNAsequences by fluorescence emission via fluorophores may occur throughprimer extension of a probe as a result of using labeled nucleotides,through molecular beacon or similar fluorophore, and through quencherbased primers or other means to detect fluorescence. For example, afirst DNA primer may be labeled with a fluorophore that absorbs light ata wavelength of 494 nanometers (nm) and fluoresces at a wavelength of521 nm (e.g. DY495), a second DNA primer may be labeled with afluorophore that absorbs light at a wavelength of 515 nm and fluorescesat a wavelength of 650 nm (e.g. DY481-XL), a third DNA primer may belabeled with a fluorophore that absorbs light at a wavelength of 559 nmand fluoresces at a wavelength of 578 nm (e.g. DY560), and a fourth DNAprimer may be labeled with a fluorophore that absorbs light at awavelength of 647 nm and fluoresces at a wavelength of 670 nm (e.g.DY636). Each of the first, second, third, and fourth DNA primers residein each of the at least four PCR tubes of a 0.2 ml volume having aplurality of DNA samples. For example, a first PCR tube contains a firstDNA sample and the first, second, third, and fourth DNA primers, asecond PCR tube contains a second DNA sample and the first, second,third, and fourth DNA primers, a third PCR tube contains the third DNAsample and the first, second, third, and fourth DNA primers, and afourth PCR tube contains a fourth DNA sample and the first, second,third, and fourth DNA primers.

In 501, the wavelength scanning apparatus 400 is initialized todetermine background fluorescence for each of the at least fourdifferent DNA samples, where each of the at least four different DNAsamples includes at least one fluorophore. Each of the different DNAsamples may contain from one to four fluorophores for detection. Each ofthe different DNA samples also may contain at least four fluorophores.Thus, during 501 the wavelength scanning apparatus 400 determines theamount and/or wavelength of fluorescence emission produced from each DNAsample that is not in response to a desired analyte. For example, inmolecular beacon fluorescence, before initiation of the PCR reactionwhich binds a primer to a DNA sequence of interest, the fluorophore isbound by a quencher and therefore will not produce a recordablefluorescent emission wavelength indicative of the presence of the DNAsequence of interest. Initialization may occur for each DNA samplewherein a light source is turned on for a period of time (e.g. 5seconds) to emit light of a first wavelength that travels through anexcitation activation aperture, an excitation aperture, an excitationlight filter, the DNA sample, a fluorescence emission dischargeaperture, an emission aperture, and a fluorescence emission light filteruntil it reaches a photodetector configured for detecting light of asecond wavelength that corresponds to the first wavelength.Initialization of each DNA sample occurs when a DNA sample is in thesample well 101 where the excitation activation aperture 102 issubstantially aligned with the excitation aperture 105 and thefluorescence emission discharge aperture 103 is substantially alignedwith the emission aperture 108 of the analysis scanner 300. For example,in the fourth position the first, second, third, and fourth DNA samplesare initialized simultaneously or nearly simultaneously. The initialreading of the wavelength by the photodetector for a DNA primer in a DNAsample is read by and stored in a computer program to determine theamount of background fluorescence of the wavelength in a DNA sample.

In 502, the biological reaction is initiated. The biological reactionmay be the amplification of DNA using polymerase chain retain (PCR) orany other isothermal amplification method compatible with the sample andthe analysis. Initiation 502 may include raising and lowering thetemperature of the heating block 100 to predetermined temperatures wherethe DNA primers will anneal to the corresponding DNA sequence andamplify by PCR or other amplification method. Annealing of the DNAprimers to the corresponding DNA sequences unquenches the fluorophore byseparation of the fluorophore and quencher such that the fluorophore mayproduce a recordable fluorescent emission wavelength.

In 503, the analysis scanner 300 is moved a position. At each position 1through 7 for a wavelength scanning apparatus 400 with four of thesample wells 101, the excitation activation aperture 102 issubstantially aligned with the excitation aperture 105 and thefluorescence emission discharge aperture 103 is substantially alignedwith the emission aperture 108 of the analysis scanner 300. For example,in the fourth position, each of the four excitation activation aperturesare substantially aligned with each of the four excitation apertures andeach of the four fluorescence emission discharge apertures aresubstantially aligned with each of the four emission apertures.

In 504, each DNA sample is analyzed for the presence and optionally thequantity of a plurality of DNA sequences by detection of the desiredfluorescent emission wavelengths. Analysis may occur at positions 1through 7 wherein a plurality of light sources are turned on for aperiod of time (e.g. 5 seconds) to emit light of a first excitationwavelength that travels through an excitation activation aperture, anexcitation aperture, an excitation light filter, and a DNA sample toexcite a fluorophore. The resulting fluorescent emission wavelength thentravels to a fluorescence emission discharge aperture, an emissionaperture, and a fluorescence emission light filter until it reaches aphotodetector configured for detecting light of a second emissionwavelength. For example, in the fourth position detection of the first,second, third, and fourth DNA samples are tested simultaneously ornearly simultaneously, each at a different excitation wavelength. Thedetection reading of the wavelength by the photodetector is read by andstored by a computer program to determine the presence, absence, and/orquantity of a DNA sequence in a DNA sample.

In 505, the wavelength scanning apparatus 400 reports the presence,absence, and/or quantity of the selected DNA sequence/s in each DNAsample. This information may be displayed, stored, transmitted, orotherwise processed. Steps 503, 504, and 505 may be repeated at eachposition 1 through position 7.

It is to be noted that the foregoing described embodiments may beconveniently implemented using conventional general purpose digitalcomputers programmed according to the teachings of the presentspecification, as will be apparent to those skilled in the computer art.Appropriate software coding may readily be prepared by skilledprogrammers based on the teachings of the present disclosure, as will beapparent to those skilled in the software art.

It is to be understood that the embodiments described herein may beconveniently implemented in forms of a software package. Such a softwarepackage may be a computer program product which employs a non-transitorycomputer-readable storage medium including stored computer code which isused to program a computer to perform the disclosed functions andprocesses disclosed herein. The non-transitory computer-readable storagemedium may include, but is not limited to, any type of conventionalfloppy disk, optical disk, CD-ROM, magnetic disk, hard disk drive,magneto-optical disk, ROM, RAM, EPROM, EEPROM, magnetic or optical card,or any other suitable non-transitory media for storing electronicinstructions.

A method 1000 is used to analyze at least four nucleic acid (e.g. DNA orribonucleic acid (RNA)) samples for at least four different nucleic acidsequences (e.g. nucleotides or oligonucleotides). A nucleic acid samplemay contain all, some, or none of the nucleic acid sequences. Eachnucleic acid sample includes a plurality of nucleic acid primers todetect the nucleic acid sequences that are present in each sample. Eachnucleic acid primer could be labeled with a fluorophore, eachfluorophore having unique absorption and emission properties. Detectionof the nucleic acid sequences by fluorescence emission via fluorophoresmay occur through primer extension of a probe as a result of usinglabeled nucleotides, through molecular beacon or similar fluorophore,and through quencher based primers or other means to detectfluorescence.

For example, a first nucleic acid primer may be labeled with afluorophore that absorbs light at a wavelength of 500 nanometers (nm)and fluoresces at a wavelength of 520 nm (e.g. Atto 488), a secondnucleic acid primer may be labeled with a fluorophore that absorbs lightat a wavelength of 564 nm and fluoresces at a wavelength of 590 nm (e.g.Atto 565), a third nucleic acid primer may be labeled with a fluorophorethat absorbs light at a wavelength of 645 nm and fluoresces at awavelength of 671 nm (e.g. DY636), and a fourth nucleic acid primer maybe labeled with a fluorophore that absorbs light at a wavelength of 747nm and fluoresces at a wavelength of 776 nm (e.g. DY750). Each of thefirst, second, third, and fourth nucleic acid primers reside in each ofthe at least four PCR tubes of a 0.2 ml volume having a plurality ofnucleic acid samples. For example, a first PCR tube contains a firstnucleic acid sample and the first, second, third, and fourth nucleicacid primers, a second PCR tube contains a second nucleic acid sampleand the first, second, third, and fourth nucleic acid primers, a thirdPCR tube contains the third nucleic acid sample and the first, second,third, and fourth nucleic acid primers, and a fourth PCR tube contains afourth nucleic acid sample and the first, second, third, and fourthnucleic acid primers.

In 1001, the wavelength scanning apparatus 400 is initialized todetermine background fluorescence for each of the at least fourdifferent nucleic acid samples, where each of the at least fourdifferent nucleic acid samples includes at least one fluorophore. Eachof the different nucleic acid samples may contain from one to fourfluorophores for detection. Each of the different nucleic samples alsomay contain at least four fluorophores. Thus, during 1001 the wavelengthscanning apparatus 400 determines the amount (e.g. intensity) of thewavelength of fluorescence emission produced from each nucleic acidsample that is not in response to a desired analyte. For example, inmolecular beacon fluorescence, before initiation of the PCR reactionwhich binds a primer to a nucleic acid sequence of interest, thefluorophore is bound by a quencher and therefore will not produce arecordable fluorescent emission wavelength indicative of the presence ofthe nucleic acid sequence of interest. Initialization may occur for eachnucleic acid sample wherein a light source is turned on for a period oftime (e.g. from 0.5 to 5 seconds) to emit light of a first wavelengththat travels through an excitation activation aperture, an excitationaperture, an excitation light filter, the nucleic acid sample, afluorescence emission discharge aperture, an emission aperture, and afluorescence emission light filter until it reaches a photodetectorconfigured for detecting light (e.g. photons) of a second wavelengththat corresponds to the first wavelength. Initialization of each nucleicacid sample occurs when a nucleic acid sample is in the sample well 101where the excitation activation aperture 102 is substantially alignedwith the excitation aperture 105 and the fluorescence emission dischargeaperture 103 is substantially aligned with the emission aperture 108 ofthe analysis scanner 300. For example, in the fourth position the first,second, third, and fourth nucleic acid samples are initializedsimultaneously or nearly simultaneously. The initial reading of thewavelength by the photodetector for a nucleic acid primer in a nucleicacid sample is read by and stored in a computer program to determine theamount of background fluorescence of the wavelength in a nucleic acidsample.

In 1002, the biological reaction is initiated. The biological reactionmay be the amplification of nucleic acid using polymerase chain retain(PCR), reverse transcription PCR, or any other isothermal amplificationmethod compatible with the sample and the analysis. Initiation 1002 mayinclude raising and lowering the temperature of the heating block 100 topredetermined temperatures where the nucleic acid primers will anneal tothe corresponding nucleic acid sequence and amplify by PCR or otheramplification method. Annealing of the nucleic acid primers to thecorresponding nucleic acid sequences unquenches the fluorophore byseparation of the fluorophore and quencher such that the fluorophore mayproduce a recordable fluorescent emission wavelength.

In 1003, the analysis scanner 300 is moved to a position. At eachposition 1 through 7 for a wavelength scanning apparatus 400 with fourof the sample wells 101, the excitation activation aperture 102 issubstantially aligned with the excitation aperture 105 and thefluorescence emission discharge aperture 103 is substantially alignedwith the emission aperture 108 of the analysis scanner 300. For example,in the fourth position, each of the four excitation activation aperturesare substantially aligned with each of the four excitation apertures andeach of the four fluorescence emission discharge apertures aresubstantially aligned with each of the four emission apertures. Theanalysis scanner 300 may be moved to the position by a controller.

In 1004, each nucleic acid sample is analyzed for the presence andoptionally the quantity of a plurality of nucleic acid sequences bydetection of the desired fluorescent emission wavelengths. Analysis mayoccur at positions 1 through 7 wherein a plurality of light sources areturned on for a period of time (e.g. from 0.5 to 5 seconds) to emitlight of a first excitation wavelength that travels through anexcitation activation aperture, an excitation aperture, an excitationlight filter, and a nucleic acid sample to excite a fluorophore. Theresulting fluorescent emission wavelength then travels to a fluorescenceemission discharge aperture, an emission aperture, and a fluorescenceemission light filter until it reaches a photodetector configured fordetecting light (e.g. photons) of a second emission wavelength. Forexample, in the fourth position detection of the first, second, third,and fourth nucleic acid samples are tested simultaneously or nearlysimultaneously, each at a different excitation wavelength. The detectionreading of the wavelength by the photodetector is read by and stored bya computer program to determine the presence, absence, and/or quantityof a nucleic acid sequence in a nucleic acid sample.

In 1005, the wavelength scanning apparatus 400 reports the presence,absence, and/or quantity of the selected nucleic acid sequence/s in eachnucleic acid sample. This information may be displayed, stored,transmitted, or otherwise processed. Steps 503, 504, and 505 may berepeated at each position 1 through position 7.

In 1006, the wavelength scanning apparatus resets to align the analysisscanner. The reset includes the analysis scanner moving to a resetposition where the analysis scanner contacts the reset trigger. Thereset trigger activates the reset button to align the analysis scannerwith the fluorescence emission discharge aperture, which ensures thesame alignment for each fluorescence detection analysis. The controllermay be configured to cause the analysis scanner to move to the resetposition.

As used herein, the term “simultaneously or nearly simultaneously” meansthat while detection of multiple wavelength fluorescence at a particularposition (e.g. position 4) has occurred simultaneously, particularfluorophores have distinct quantum efficiencies, such that fluorescenceoccurs at a different time (e.g. milliseconds) as compared to anotherparticular fluorophore.

Although the subject matter has been described in language specific tostructural features and/or methodological acts, it is to be understoodthat the subject matter defined in the appended claims is notnecessarily limited to the specific features or acts described above.Rather, the specific features and acts described above are disclosed asexample forms of implementing the claims.

It is believed that the present invention and many of its attendantadvantages will be understood from the foregoing description. It is alsobelieved that it will be apparent that various changes may be made inthe form, construction and arrangement of the components thereof withoutdeparting from the scope and spirit of the invention or withoutsacrificing all of its material advantages. The form herein beforedescribed being merely an explanatory embodiment thereof, it is theintention of the following claims to encompass and include such changes.

1. A wavelength scanning apparatus for measuring fluorescence emissionof a plurality of nucleic acid samples, comprising: a structuralsupport, the structural support comprising a reset button and a resettrigger, wherein the reset button is in mechanical communication withthe reset trigger; a heating block attached to the structural support,the heating block comprising a plurality of sample wells, wherein eachof the plurality of sample wells forms an opening on a top side of theheating block, a plurality of excitation activation apertures, and aplurality of fluorescence discharge apertures, wherein each of theplurality of sample wells transitions to a corresponding excitationactivation aperture of the plurality of excitation activation aperturesand a corresponding fluorescence discharge aperture of the plurality ofexcitation activation apertures, and wherein each of the plurality ofexcitation activation apertures forms an opening on a bottom side of theheating block and each of the plurality of fluorescence dischargeapertures forms an opening on a front side of the heating block; astepper motor attached to the structural support; an analysis scannerattached to the stepper motor, where the stepper motor moves theanalysis scanner laterally, wherein the analysis scanner comprises aflat surface, the flat surface comprising a plurality of excitationapertures, where each of the plurality of excitation apertures comprisesa top and a bottom, and where each of the plurality of excitationapertures comprises an excitation light filter; a front wall, the frontwall having a plurality of emission apertures, where each of theplurality of emission apertures comprises a back and a front, and whereeach of the plurality of emission apertures comprises a fluorescenceemission light filter; a plurality of light sources, where each of theplurality of light sources is disposed in a corresponding excitationaperture of the plurality of excitation apertures; a plurality ofphotodetectors, where each of the plurality of photodetectors isdisposed in a corresponding emission aperture of the plurality ofemission apertures; and where the flat surface of the analysis scannersubstantially aligns with the bottom side of the heating block and thefront wall of the analysis scanner substantially aligns with the frontside of the heating block.
 2. The wavelength scanning apparatus of claim1, wherein the fluorescence emission discharge aperture of the heatingblock is from 0.5 to 3.0 millimeters from the top of the excitationactivation aperture.
 3. The wavelength scanning apparatus of claim 2,wherein the fluorescence emission discharge aperture of the heatingblock is cylindrical having a diameter of 2 millimeters.
 4. A nucleicacid analysis method for performing fluorescence detection of multiplefluorescence emission wavelengths simultaneously or nearlysimultaneously, comprising: initializing at least four photodetectorswith a plurality nucleic acid samples, wherein each nucleic acid samplesincludes a plurality of unreacted nucleic acid primers, where eachnucleic acid primer has a fluorophore; reacting the plurality of nucleicacid primers with a plurality of nucleic acid sequences to measure thefluorophores of the plurality of nucleic acid primers; moving ananalysis scanner to a position to substantially align a plurality ofexcitation activation apertures with a plurality of excitation aperturesand a plurality of fluorescence discharge apertures with a plurality ofemission apertures where light from a plurality of light sources excitesthe plurality of fluorophores in the plurality of nucleic acid samples;analyzing the nucleic acid samples for a plurality of fluorescentemission wavelengths, wherein the fluorescent emission wavelengthscomprise photons of each of the plurality of fluorescent emissionwavelengths; and reporting the readings from a plurality ofphotodetectors that measure the fluorescent emission wavelengths of thenucleic acid samples.
 5. The method of claim 4, further comprisingresetting the analysis scanner after reporting the readings from aplurality of photodetectors that measure the fluorescent emissionwavelengths of the nucleic acid samples.
 6. The method of claim 5,wherein the analysis scanner is reset by a controller.
 7. The method ofclaim 6, wherein the controller is configured to move the analysisscanner to contact a reset trigger.
 8. The method of claim 6, whereinthe analysis scanner is moved by the controller.
 9. The method of claim4, further comprising repeating the moving, the analyzing, and thereporting when the plurality of nucleic acid samples contains unanalyzedfluorophores.
 10. The method of claim 4, where the plurality of nucleicacid samples is at least three.
 11. The method of claim 4, where theplurality of nucleic acid samples is at least four.
 12. The method ofclaim 4, where the nucleic acid sample is a nucleic acid sample.
 13. Themethod of claim 4, where the nucleic acid sample is an RNA sample.
 14. Anucleic acid analysis method for performing fluorescence detection ofmultiple fluorescence emission wavelengths simultaneously or nearlysimultaneously, comprising: initializing a first photodetector and asecond photodetector with at least a first quenched fluorophore and asecond quenched fluorophore; reacting a first nucleic acid sequence tounquench the first fluorophore to provide a first unquenchedfluorophore, and a second nucleic acid sequence to unquench the secondfluorophore to provide a second unquenched fluorophore, wherein a firstnucleic acid sample contains the first nucleic acid sequence and thesecond nucleic acid sequence, and a second nucleic acid sample containsthe first nucleic acid sequence and the second nucleic acid sequence;moving an analysis scanner to a position to substantially align a firstexcitation activation aperture with a first excitation aperture and afirst fluorescence discharge aperture with a first emission aperturewhere a first light source is capable of exciting the first unquenchedfluorophore in the first nucleic acid sample, and a second excitationactivation aperture with a second excitation aperture and a secondfluorescence discharge aperture with a second emission aperture where asecond light source is capable of exciting the second unquenchedfluorophore in the second nucleic acid sample; analyzing a firstfluorescent emission wavelength by a first photodetector for a firstfluorescent emission wavelength, and analyzing a second fluorescentemission wavelength by a second photodetector for a second fluorescentemission wavelength; and reporting the readings from the firstphotodetector and the second photodetector to detect the firstfluorescent emission wavelength and the second fluorescent emissionwavelengths nearly simultaneously.
 15. The method of claim 14, furthercomprising initializing a third photodetector with a third quenchedfluorophore; reacting a third nucleic acid sequence to unquench thethird fluorophore to provide a third unquenched fluorophore, wherein thefirst nucleic acid sample further contains the third nucleic acidsequence, the second nucleic acid sample further contains the thirdnucleic acid sequence, and a third nucleic acid sample contains thefirst nucleic acid sequence, the second nucleic acid sequence, and thethird nucleic acid sequence; moving the analysis scanner to a positionto substantially align a third excitation activation aperture with athird excitation aperture and a third fluorescence discharge aperturewith a third emission aperture where a third light source is capable ofexciting the third unquenched fluorophore in the third nucleic acidsample; analyzing a third fluorescent emission wavelength by a thirdphotodetector for a third fluorescent emission wavelength; and reportingthe readings from the first photodetector, the second photodetector, andthe third photodetector to detect the first fluorescent emissionwavelength, the second fluorescent emission wavelength, and the thirdfluorescent emission wavelength nearly simultaneously.
 16. The method ofclaim 15, further comprising initializing a fourth photodetector with afourth quenched fluorophore; reacting a fourth nucleic acid sequence tounquench the fourth fluorophore to provide a fourth unquenchedfluorophore, wherein the first nucleic acid sample further contains thefourth nucleic acid sequence, the second nucleic acid sample furthercontains the fourth nucleic acid sequence, the third nucleic acid samplefurther contains the fourth nucleic acid sequence, and a fourth nucleicacid sample contains the first nucleic acid sequence, the second nucleicacid sequence, the third nucleic acid sequence, and the fourth nucleicacid sequence; moving the analysis scanner to a position tosubstantially align a fourth excitation activation aperture with afourth excitation aperture and a fourth fluorescence discharge aperturewith a fourth emission aperture where a fourth light source is capableof exciting the fourth unquenched fluorophore in the fourth nucleic acidsample; analyzing a fourth fluorescent emission wavelength by a fourthphotodetector for a fourth fluorescent emission wavelength; andreporting the readings from the first photodetector, the secondphotodetector, the third photodetector, and the fourth photodetector todetect the first fluorescent emission wavelength, the second fluorescentemission wavelength, the third fluorescent emission wavelength, and thefourth fluorescent emission wavelength nearly simultaneously.
 17. Themethod of claim 16, further comprising repeating the moving, theanalyzing, and the reporting when the first, second, third, and fourthnucleic acid samples contain unanalyzed fluorophores.